Technical Field
The present disclosure relates to a confocal scanner configured to obtain a super resolution by optical processing, and more particularly, to a confocal scanner capable of reducing complication of a positioning operation of an optical component and a confocal microscope using the confocal scanner.
Related Art
A confocal microscope configured to irradiate a light to an object with pinpoint, to selectively detect only a focusing light from an irradiation point, to scan the irradiation point and to obtain an image is an optical microscope capable of reconstructing an image of a high resolution and three-dimensional information. The confocal microscope is widely used in the biological science field and the like, and a variety of related technologies have been suggested.
For example, Patent Document 1 discloses a confocal scanner, which is a scan unit of a confocal microscope, includes a micro lens disk having a plurality of micro lenses and a pinhole disk having pinholes formed in the same pattern as the micro lenses, and is configured to perform a multi-beam scanning by rotating the disks while irradiating an illumination light.
In the related art, an optical microscope including the confocal microscope has an Abbe's diffraction limit, i.e., a resolution limit based on a theory that an object smaller than a half wavelength of the light to be irradiated to the object cannot be seen. In recent years, a super resolution technology of obtaining an image having a resolution beyond the resolution limit has been developed and put to practical use.
For example, Patent Document 2 discloses a technology of using a scan mask, which is configured to modulate spatial intensity distributions of an excitation light from a light source and a return light from an object, to capture an image having a high frequency component beyond a resolution limit, and performing high frequency enhancement processing to obtain a super resolution. Also, Non-Patent Document 1 discloses a technology of using a shutter, which is configured to pass an illumination light from a light source apparatus in a strobe shape, to capture hundreds of images having a plurality of bright spots recorded therein while slightly changing positions of the bright spots, performing image processing in which the bright spot becomes a half size for each image, and synthesizing the respective images to obtain a super resolution image.
According to the super resolution technologies disclosed in Patent Document 2 and Non-Patent Document 1, since the complicated or the large amount of image processing is performed, the calculation load is high and much time is consumed for the processing. That is, the corresponding super resolution technologies are inappropriate to the real-time observation.
In contrast, Patent Document 3 discloses a technology capable of obtaining a super resolution image at high speed by optical processing. FIG. 19 shows a configuration of an optical system of a microscope system disclosed in Patent Document 3.
As shown in FIG. 19, a microscope system 400 is configured to irradiate a laser light, which is to be emitted from a light source 410, to a sample 430, and to capture a return light with a camera 420. The collimated illumination light emitted from the light source 410 is divided into a plurality of illumination light beamlets by a micro lens array 411. The illumination light beamlets are reflected on a galvanometer mirror 413, and are concentrated on the sample 430 by an objective lens 414.
The sample 430 is configured to radiate the return light based on the illumination light. In particular, in case of fluorescent sample observation, the sample 430 is a specific structure dyed by a fluorescent dye and the like, and is configured so that fluorescent dye molecules are excited by the illumination light and radiate the fluorescence having a longer wavelength than the illumination light.
The return light from the sample 430 is reflected on the galvanometer mirror 413, is reflected on a beam splitter 412 and then passes through a lens 415. The return light having passed through the lens 415 reaches a pinhole array 416 having a plurality of pinholes. However, only the light from a focal plane of the sample 430 is focused on the pinhole array 416 and passes through the pinholes.
The return light having passed through the pinhole passes through a micro lens array 417 and a micro lens array 418 each of which having a plurality of micro lenses, is reflected on the galvanometer mirror 413, and is captured by the camera 420. The return light is a part of the sample 430 on which the illumination light beamlets are reflected, but can scan the entire sample 430 by changing a direction of the galvanometer mirror 413.
Here, the pinhole array 416 is precisely arranged so that each pinhole is disposed at a position conjugate with a focusing spot of the objective lens 414. Also, each pinhole of the pinhole array 416 is precisely arranged at a focal position of each micro lens of the micro lens array 417. Further, each micro lens of the micro lens array 417 and each micro lens of the micro lens array 418 are precisely arranged to be coaxial with each other.
A focal length of each micro lens of the micro lens array 418 is set to be shorter than a focal length of each micro lens of the micro lens array 417.
In the above configuration, the return light of the focusing plane having passed through the pinholes of the pinhole array 416 is converted into a parallel light by the micro lens array 417, which is then incident to the micro lens array 418. Since the focal length of each micro lens of the micro lens array 418 is shorter than the focal length of each micro lens of the micro lens array 417, a numerical aperture of the return light increases when passing through the micro lens array 417. For example, when the focal length of each micro lens of the micro lens array 418 is a half of the focal length of each micro lens of the micro lens array 417, the return light is converted into a light beam having a double numerical aperture.
When the light beam is captured by the camera 420 while changing a direction of the galvanometer mirror 413, a super resolution image of the sample 430 can be obtained. At this time, since it is not necessary to perform the troublesome image processing and the plurality of capturing processing, it is possible to simply obtain the super resolution image at high speed.    [Patent Document 1] Japanese Patent Application Publication No. Hei 10-062691A    [Patent Document 2] Japanese Patent Application Publication No. 2012-78408A    [Patent Document 3] International Patent Application Publication No. WO2013/126762    [Non-Patent Document 1] Schulz, O. et al. Resolution doubling in fluorescence microscopy with confocal spinning-disk image scanning microscopy. Proceedings of the National Academy of Sciences of United States of America, Vol. 110, pp. 21000-21005 (2013)
As described above, it is possible to simply obtain the super resolution image at high speed by using the confocal scanner configured to obtain a super resolution with the optical processing. However, in case of the confocal scanner configured to obtain a super resolution image with the optical processing, it is necessary to perform the precise positioning with respect to each optical component of the micro lens array for the illumination light, the objective lens, the two micro lens arrays for the return light, and the pinhole array. According to the optical system disclosed in Patent Document 3, the optical components are independently arranged with being spatially spaced, so that the precise positioning is not easy.
The precise positioning of the optical component causes the cost increase and is easily influenced by environment and temporal changes, and the maintenance thereof is complicated. Therefore, it is preferably to reduce a burden on the precise positioning as much as possible.